Kupffer cells are fixed macrophages that line the hepatic sinusoids. These cells serve as a selective filter to remove particulate matter from the portal circulation and to defend the hepatic parenchyma against endotoxin. Following liver injury, Kupffer cells become "activated". They display enhanced phagocytosis, chemotaxis, cytotoxicity and release reactive mediators. Although the function of Kupffer cells and their biological mediators in normal systemic host defense has been established, their potential role in chemical induced hepatic injury is unknown. We have been studying the hepatotoxic effects of the analgesic acetaminophen. Using isolated perfused rat liver, we observed an inhibition of hepatic respiration and selective pericentral necrosis 24 hrs following treatment of fasted rats with acetaminophen. Interestingly, in fed rats, although there were no apparent signs of necrosis, we did observe a large infiltration of mononuclear cells into the pericentral region of the liver lobule. It is our hypothesis that the mononuclear cell infiltrate is composed of newly recruited and "activated" Kupffer cells, and that these cells contribute to hepatocellular injury caused by acetaminophen. The overall objective of this proposal is to characterize the mechanism by which "activated" Kupffer cells accumulate in the liver following acetaminophen treatment, and to elucidate their potential role in hepatotoxicity. For these studies, we will use isolated hepatocytes and Kupffer cells maintained in culture in conjunction with isolated perfused liver. In preliminary experiments we found that conditioned medium from hepatocytes treated with acetaminophen, but not acetaminophen itself, induced Kupffer cell chemotaxis and superoxide anion release and stimulated Kupffer cell phagocytosis. This suggests that acetaminophen injured hepatocytes release factors that attract and "activate" macrophages. We will use biophysical and biochemical techniques to characterize these factors and their roles in chemical induced liver injury.